ABSTRACT There are no effective therapies to treat Cryptosporidium, an important opportunistic pathogen of AIDS patients that is now recognized as significant cause of diarrheal disease worldwide. Individuals who are immunocompromised are highly susceptible to cryptosporidiosis and experience chronic, debilitating and sometimes fatal diarrhea unless the immune system can be reconstituted. Development of effective therapeutics for cryptosporidiosis is undoubtedly a medical imperative. Many therapeutics in clinical use today are, or are derived from, natural products and a large untapped source of naturally occurring bioactive compounds is the ocean. We propose to search for novel anti-cryptosporidial compounds with therapeutic potential in a unique chemical library of marine compounds. The Harbor Branch Oceanographic Institute (HBOI) natural products library consists of 125 purified compounds and 6500 highly enriched fractions from extracts of marine organisms collected from a wide range of habitats including deep sea enviroments; the diversity and novelty of the compounds contained in this library has been well documented and includes compounds effective against the related apicomplexan pathogen Plasmodium falciparum. To illustrate the potential success of this approach, in screening just a few strains of marine symbiotic bacteria for bioactive compounds we discovered the compound Tartrolon D that is broadly effective at nM levels against multiple apicomplexan parasites including Cryptosporidium and Plasmodium. Therefore we hypotheisze that screening the HBOI library against Cryptosporidium will identify multiple novel compounds with potent activity against this parasite. We will test this hypothesis through completion of the following aims: Aim 1: Screen the HBOI Peak library against Cryptosporidium parvum-infected cells using a high throughput assay for cell viability. The library will be tested against Cryptosporidium-infected intestinal epithelial cells and anti-parasitic activity detected by an increase in host-cell viability. Aim 2: Prioritization of anti-Cryptosporidium active fractions and purified compounds. Positive hits will be verified by examining activity against luciferase-expressing Cryptosporidium in standard cell culture and in intestinal and lung organoids. Positive hits will also be tested against Toxoplasma gondii. Aim 3. Isolation and structure elucidation of active compounds: Positive fractions with the greatest potential for discovery of novel inhibitors will be taken through further purification and the structures of the active compounds defined using spectroscopic methods. Currently there are only a handful of anti-cryptosporidial compounds in various stages of development. Expanding drug discovery, especially into the realm of natural products, is critical to the ultimate goal of bringing an effective anti-Cryptosporidium therapeutic to the clinics.